While a graft-versus-leukemia (GVL) effect has been established in patients who undergo hematopoietic stem cell transplant (SCT), suggesting acute lymphoblastic leukemia (ALL) may be controlled by cellular immune-mediated pathways, the relative lack of efficacy of donor lymphocyte infusion for ALL suggests that leukemic cells are poorly immunogenic. New methods that can overcome poor tumor immunogenicity and have the potential to be efficacious for treatment of ALL with less toxicity than standard approaches used to treat high risk and relapsed disease, including SCT, need to be pursued (Horowitz, et al., 1990, Blood 75(3):555-562; Mehta, 1993, Leuk Lymphoma 10(6):427-432).
Chimeric antigen receptors (CAR) are molecules combining antibody-based specificity for tumor-associated surface antigens with T cell receptor-activating intracellular domains with specific anti-tumor cellular immune activity (Eshhar, 1997, Cancer Immunol Immunother 45(3-4) 131-136; Eshhar et al., 1993, Proc Natl Acad Sci USA 90(2):720-724; Brocker and Karjalainen, 1998, Adv Immunol 68:257-269). These CARs allow a T cell to achieve MHC-independent primary activation through single chain Fv (scFv) antigen-specific extracellular regions fused to intracellular domains that provide T cell activation and co-stimulatory signals. Second and third generation CARs also provide appropriate co-stimulatory signals via CD28 and/or CD137 (4-1BB) intracellular activation motifs, which augment cytokine secretion and anti-tumor activity in a variety of solid tumor and leukemia models (Pinthus, et al, 2004, J Clin Invest 114(12):1774-1781; Milone, et al., 2009, Mol Ther 17(8):1453-1464; Sadelain, et al., 2009, Curr Opin Immunol 21(2):215-223).
Most investigators have acheived efficient CAR gene transfer of human tumor and HIV antigens into human T cells via retrovirus or HIV-derived lentivirus, and some of these cell therapy products have advanced to Phase I/II trials (Deeks et al., 2002, Mol Ther 5(6):788-797; Kershaw, et al., 2006, Clin Cancer Res 12(20 Pt 1):6106-6115; Pule, et al., 2008, Nat Med 14(11):1264-1270; Till, et al., 2008, Blood 112(6):2261-2271). Recently, the use of CD19-targeted CAR+T cells in three patients with CLL has been reported (Porter et al., 2011, N Eng J Med, 365: 725-733). Two of three of these patients with refractory disease and high tumor burdens entered a complete remission after 4 weeks. These responses have been sustained and the CAR+T cells persisted for >6 months, suggesting the efficacy of this technology. Approaches using integrating viral vectors have clear advantages, including long-term expression of the CAR on infused cells across multiple cell divisions. However, iterative clinical trials which rapidly incorporate CAR design innovations may be difficult to implement using viral vectors, because of the complexity of release testing and the high expense of vector production. In addition, there are regulatory concerns using this approach. This has clearly been seen in the case of a retroviral vector used in gene modification of hematopoietic stem cells in the treatment of X-linked severe combined immunodeficiency (Hacein-Bey-Abina et al., 2008, J Clin Invest 118(9):3132-3142). In the case of lentiviral vectors, or in the setting of gene modification of mature lymphocytes, this is a theoretical concern, but it is an issue for regulators of gene and cell therapy technologies.
Electroporation-mediated mRNA transfection is a potentially complementary approach for gene expression that does not result in permanent genetic modification of cells. The use of mRNA for gene therapy applications was first described by Malone et al. in the context of liposome-mediated transfection (Malone, et al., 1989, Proc Natl Acad Sci USA 86(16):6077-6081). Successful electroporation of mRNA into primary T lymphocytes has now been developed and used for efficient TCR gene transfer (Zhao, et al., 2006, Mol Ther 13(1):151-159; Zhao, et al., 2005, J. Immunol. 174(7):4415-4423). More recently, CARs directed against the Her2/neu antigen were introduced into T cells by mRNA electroporation and were found to be more effective than Her2/neu antibodies in a breast cancer xenograft model (Yoon, et al., 2009, Cancer Gene Ther 16(6):489-497). Other human target antigen-directed CARs introduced into T cells by mRNA electroporation include those targeting CEA and ErbB2 (Birkholz et al., 2009, Gene Ther 16(5):596-604). While a number of articles report efficacy using this approach in solid tumors after intratumoral injection or in local injection intraperitoneal models, similar success has not been demonstrated in disseminated leukemia pre-clinical models possibly due to the difficulty in achieving efficacy in a disseminated model using a transient expression system (Rabinovich, et al., 2009, Hum Gene Ther 20(1):51-61).
CD19 is a surface antigen restricted to B cells, and is expressed on early pre-B cells and a majority of B cell leukemias and lymphomas (Nadler, et al., 1983 J Immunol 131(1):244-250). This makes CD19 an attractive antigen for targeted therapy as it is expressed on the malignant cell lineage and a specific subset of early and mature B lymphocytes but not hematopoietic stem cells. It has been postulated that CD19 depletion allows for eventual restoration of a normal B cell pool from the CD19 negative precursor population (Cheadle et al., 2010, J Immunol 184(4):1885-1896). Experience with rituximab, the anti-CD20 monoclonal antibody used for treatment of B cell malignancies and autoimmune disorders, has shown that therapy induced B cell deficiency is well tolerated (Plosker and Figgitt, 2003, Drugs 63(8):803-843; van Vollenhoven, et al., 2010, J Rheumatol 37(3):558-567).
Adoptive transfer of CTLs has shown great promise in both viral infections and cancers. After many years of disappointing results with chimeric antigen receptor (CAR) T-cell therapy, improved culture systems and cell engineering technologies are leading to CAR T cells with more potent antitumor effects (Sadelain et al., 2009, Curr Opin Immunol 21:215-23). Results from recent clinical trials indicate improved clinical results with CARs introduced with retroviral vectors (Till et al., 2008, Blood 112:2261-71; Pule et al., 2008, Nat Med 14:1264-70). Perhaps not surprisingly, these CAR T cells also exhibit enhanced toxicity (Brentjens et al., 2010, Mol Ther 18:666-8; Morgan et al., 2010, Mol Ther 18:843-51). Recent editorials have discussed the need for safer CARs (Heslop, 2010, Mol Ther 18:661-2; Buning et al., 2010, Hum Gene Ther 21:1039-42).
Thus, there is an urgent need in the art for compositions and methods for providing additional compositions and methods to affect adoptive transfer of CTLs. The present invention addresses this need.